The long term objective and specific aim of this proposal is to perfect a technique so that the state-of-the-art cell kinetic tool, flow cytometry, can be used to construct cell kinetic profiles of various states of human corneal epithelium: 1) normal, 2) bullous keratopathy, 3) diabetes, 4) neurotrophic keratopathy, 5) corneal dystrophy, 6) epithelial tumors and 7) marginal ulceration due to autoimmune disease. Epithelia from these various conditions of the cornea will be obtained by medically appropriate scraping during surgical procedures. Isolated nuclear suspensions will be produced from the scrapings. The nuclear suspensions then will be stained with Vindelov's propidium iodide solution. The corneal scrapings, the preparation and staining of the samples will take place at the University of Arkansas for Medical Sciences (UAMS) in Little Rock. Samples will be analyzed on a Coulter Electronics EPICS V system at the National Center for Toxicological Research (NCTR). For each sample cell kinetic data will be stored in memory and when sufficient numbers of samples corneal condition are obtained statistical comparisons will be made between all combinations available, e.g., normal vs. diabetic, normal vs. neurotrophic keratopathy, diabetic vs. neurokeratopathy, etc. In this way a cell kinetic profile characteristic of each different condition may be obtained. These data may indicate the presence of a proliferative defect in one or more of the conditions examined. Because of the strong possibility of a circadian oscillation in cell cycle variability in human corneal epithelium, strict attention will be paid to the time when the scraping is performed. When enough numbers of samples have been accumulated/time/point/condition, statistical comparison between the DNA histograms will be made combinations of interest, e.g., normal epithelia from early morning scrapings vs. normal epithelia from early evening scrapings (2000). Statistically significant differences between 2 groups on the basis of time of day would suggest that oscillation is characteristic of the human corneal epithelium. If this pilot project demonstrates nothing else except that isolated nuclei from human corneal epithelium can be prepared, processed and analyzed by flow cytometry, a significant contribution, will have been made because the versatility and sophistication of flow cytometry then be utilized in further work on this human tissue.